Identification of a de novo LRP1 mutation in a Saudi family with Tetralogy of Fallot.

BACKGROUND: Tetralogy of Fallot (TOF) is a rare, complex congenital heart defect caused by genetic and environmental interactions that results in abnormal heart development during the early stages of pregnancy. Genetic basis of TOF in Saudi populations is not yet studied. Therefore, the objective of this study is to screen for the molecular defects causing TOF in Saudi patients. METHODS: A family with non-syndromic TOF was recruited from the Western region of Saudi Arabia. Whole exome sequencing (WES) was performed on the proband and her parents. The identified candidate variant was verified by sanger sequencing. Also, different computational biology tools were used to figure out how candidate variants affect the structure and function of candidate protein involved in TOF. RESULTS: A novel heterozygous de novo mutation in LRP1 (p. G3311D) gene was identified in the index case. Also, this variant was absent in the in-house exome sequencing data of 80 healthy Saudi individuals. This variant was predicted to be likely pathogenic, as it negatively affects the biophysical chemical properties and stability of the protein. Furthermore, functional biology data from knock out mouse models confirms that molecular defects in LRP1 gene leads to cardiac defects and lethality. This variant was not previously reported in both Arab and global population genetic databases. CONCLUSION: The findings in this study postulate that the LRP1 variant has a role in TOF pathogenesis and facilitate accurate diagnosis as well as the understanding of underlying molecular mechanisms and pathophysiology of the disease.

Exploring the role of Islam on the lived experience of patients with Long QT Syndrome in Saudi Arabia.

Genetic services are rapidly growing in the Arab world leading to increasing number of patients being diagnosed with genetic disorders. Islam is the only/major religion of the local population in these countries. Muslim patients integrate religion in virtually every aspect of their lives, and it is vital to understand the role of Islam on their coping and decision-making in the context of genetic counseling. This will help provide patients with the most appropriate services aligned to their religious beliefs and will improve outcomes. Increasing numbers of patients are being diagnosed with Long QT syndrome in Saudi Arabia. Using semi-structured interviews, this study explored the role of Islam on the lived experience of 13 Saudi participants diagnosed with autosomal dominant Long QT syndrome (3/13) or who are carriers of Jervell and Lange-Nielsen syndrome (10/13). The interviews investigated how they made sense of living with the condition in light of their religion/spirituality. The data were analyzed using interpretative phenomenological analysis and produced four superordinate themes: 1) Common belief and idiosyncratic interpretation; 2) Using religion to justify positive reframing of current illnesses; 3) Interplay between belief in medicine and in religion; and 4) Complex impact of diagnosis on religiosity. The results show that the participants' idiosyncratic interpretations of the religious principles, not the principles themselves, had an important influence on their coping, medical decision-making, perceptions regarding the cause of their disease, and compliance with medical advice. A novel insight of the current study is that the personal understanding and interpretation of medical information played the greatest role in the decision-making process, and not the religious beliefs. Thus, it is important for health professionals to give patients' information in a manner that is clear and detailed in order for them to facilitate an informed decision, and to ensure that they fully understand the implications.

Saudi Familial Hypercholesterolemia Patients With Rare LDLR Stop Gain Variant Showed Variable Clinical Phenotype and Resistance to Multiple Drug Regimen.

Familial hypercholesterolemia (FH), a well-known lipid disease caused by inherited genetic defects in cholesterol uptake and metabolism is underdiagnosed in many countries including Saudi Arabia. The present study aims to identify the molecular basis of severe clinical manifestations of FH patients from unrelated Saudi consanguineous families. Two Saudi families with multiple FH patients fulfilling the combined FH diagnostic criteria of Simon Broome Register, and the Dutch Lipid Clinic Network (DLCN) were recruited. LipidSeq, a targeted resequencing panel for monogenic dyslipidemias, was used to identify causative pathogenic mutation in these two families and in 92 unrelated FH cases. Twelve FH patients from two unrelated families were sharing a very rare, pathogenic and founder LDLR stop gain mutation i.e., c.2027delG (p.Gly676Alafs*33) in both the homozygous or heterozygous states, but not in unrelated patients. Based on the variant zygosity, a marked phenotypic heterogeneity in terms of LDL-C levels, clinical presentations and resistance to anti-lipid treatment regimen (ACE inhibitors, ß-blockers, ezetimibe, statins) of the FH patients was observed. This loss-of-function mutation is predicted to alter the free energy dynamics of the transcribed RNA, leading to its instability. Protein structural mapping has predicted that this non-sense mutation eliminates key functional domains in LDLR, which are essential for the receptor recycling and LDL particle binding. In conclusion, by combining genetics and structural bioinformatics approaches, this study identified and characterized a very rare FH causative LDLR pathogenic variant determining both clinical presentation and resistance to anti-lipid drug treatment.

Identification of a Rare Exon 19 Skipping Mutation in ALMS1 Gene in Alström Syndrome Patients From Two Unrelated Saudi Families.

Background: Alström syndrome (AS) is a very rare childhood disorder characterized by cardiomyopathy, progressive hearing loss and blindness. Inherited genetic variants of ALMS1 gene are the known molecular cause of this disease. The objective of this study was to characterize the genetic basis and understand the genotype-phenotype relationship in Saudi AS patients. Methods: Clinical phenotyping and whole-exome sequencing (WES) analysis were performed on six AS patients belonging to two unrelated consanguineous Saudi families. Sanger sequencing was performed to determine the mode of inheritance of ALMS1 variant in first-degree family relatives and also to ensure its rare prevalence in 100 healthy population controls. Results: We identified that Alström patients from both the families were sharing a very rare ALMS1, 3'-splice site acceptor (c.11873-2 A>T) variant, which skips entire exon-19 and shortens the protein by 80 amino acids. This disease variant was inherited by AS patients in autosomal recessive mode and is not yet reported in any population-specific genetic databases. AS patients carrying this mutation showed heterogeneity in clinical presentations. Computational analysis of the mutant centroid structure of ALMS1 mRNA revealed that exon-19 skipping enlarges the hairpin loop and decreases the free energy, eventually affecting its folding pattern, stability, and function. Hence, we propose c.11873-2A as an AS causative potential founder mutation in Saudi Arabia because it is found in two families lacking a common lineage. Conclusions: We conclude that WES analysis potentially helps in clinical phenotyping, early diagnosis, and better clinical management of Alström patients showing variable clinical expressivity.

Identification of Causative Variants Contributing to Nonsyndromic Orofacial Clefts Using Whole-Exome Sequencing in a Saudi Family.

Objectives: Nonsyndromic orofacial clefts (NSOFCs) are the most common craniofacial malformations observed across the globe. They are classified into three types: (a) cleft palate, (b) cleft lip, and (c) cleft lip and palate. To identify the potential candidate genes contributing to polygenic diseases such as NSOFC, linkage analyses, genome-wide association studies, and genomic rearrangements can be used. Genomic analyses, based on massively parallel next-generation sequencing technologies, play a vital role in deciphering the genetic bases of NSOFCs. Materials and Methods: In this study, whole exome sequencing was employed to detect genes that likely contributed to the NSOFC phenotype in a consanguineous Saudi family. Results: The exome analysis revealed NRP1 (rs35320960) as one potential candidate gene that is involved in bone differentiation. The RPL27A gene (rs199996172), which plays a crucial role in ribosome biogenesis, also passed all filters to serve as a candidate gene for NSOFC in this family. Rare variants are situated within the 5' UTR of these two genes. Conclusion: The study suggests that rare variants in NRP1 and RPL27A may be associated with NSOFC disease etiology.

Myocardial infarction biomarker discovery with integrated gene expression, pathways and biological networks analysis.

Myocardial infarction (MI) is the most prevalent coronary heart disease caused by the complex molecular interactions between multiple genes and environment. Here, we aim to identify potential biomarkers for the disease development and for prognosis of MI. We have used gene expression dataset (GSE66360) generated from 51 healthy controls and 49 patients experiencing acute MI and analyzed the differentially expressed genes (DEGs), protein-protein interactions (PPI), gene network-clusters to annotate the candidate pathways relevant to MI pathogenesis. Bioinformatic analysis revealed 810 DEGs. Their functional annotations have captured several MI targeting biological processes and pathways like immune response, inflammation and platelets degranulation. PPI network identify seventeen hub and bottleneck genes, whose involvement in MI was further confirmed by DisGeNET database. OpenTarget Platform reveal unique bottleneck genes as potential target for MI. Our findings identify several potential biomarkers associated with early stage MI providing a new insight into molecular mechanism underlying the disease.

A Novel Homozygous Frameshift Variant in DYM Causing Dyggve-Melchior-Clausen Syndrome in Pakistani Patients.

Background: Dyggve-Melchior-Clausen syndrome (DMC) is a skeletal dysplasia with associated defects of brain development and intelligence. The truncating pathogenic variants in DYM are the most frequent cause of DMC. Smith-McCort (SMC), another skeletal dysplasia, is also caused by non-synonymous DYM variants. Methods and Results: In the current study, we examined a Pakistani consanguineous family with three affected members. Clinical features like spondyloepimetaphyseal dysplasia, indicative of characteristic skeletal abnormalities, and intellectual disability were observed. Our male patients had microcephaly and coarse facial features while the female patient did not represent microcephaly or abnormal facies, which are significant features of DMC patients. Sanger sequencing identified a novel homozygous frameshift insertion (c.95_96insT, p.W33Lfs*14) in DYM, which likely leads to nonsense-mediated decay (NMD). Conclusion: The novel frameshift change verifies the fact that pathogenic variants in DYM are the most frequent cause of DMC.

Whole exome sequencing identifies rare biallelic ALMS1 missense and stop gain mutations in familial Alström syndrome patients.

Alström syndrome (AS, OMIM ID 203800) is a rare childhood multiorgan disorder, which is widely studied in non-Arab ethnic patients. The clinical and molecular basis of AS and the mode of disease inheritance in consanguineous Arab populations is not well investigated. Therefore, to identify the molecular basis of AS in familial forms, the present study performed whole exome sequencing of 5 AS patients belonging to 2 different Bedouin families from Saudi Arabia. The present study identified the AS causative rare biallelic mutations in ALMS gene:T376S in exon 5 and S909* in exon 8 for family A and an R2721* in exon 10 (R2721*) for family B. ALMS1 targeted genetic sequencing of healthy population controls and family members has confirmed its extremely rare frequency and autosomal recessive mode of inheritance. The truncating mutations S909* and R2721* could cause the loss of CC domains and ALMS motif on C-terminal end of the protein and creates unstable protein, which eventually undergoes intracellular degradation. The premature protein truncating mutations described in our study may eventually provide further insight into the functional domains of the ALMS1 protein and contribute to the understanding of the phenotypic spectrum of AS. Whole exome sequencing based molecular diagnosis is expected to rule out ambiguity surrounding clinical diagnosis of suspected AS cases.

Exome sequencing and metabolomic analysis of a chronic kidney disease and hearing loss patient family revealed RMND1 mutation induced sphingolipid metabolism defects.

Mitochondrial disorders (MIDs) shows overlapping clinical presentations owing to the genetic and metabolic defects of mitochondria. However, specific relationship between inherited mutations in nuclear encoded mitochondrial proteins and their functional impacts in terms of metabolic defects in patients is not yet well explored. Therefore, using high throughput whole exome sequencing (WES), we screened a chronic kidney disease (CKD) and sensorineural hearing loss (SNHL) patient, and her family members to ascertain the mode of inheritance of the mutation, and healthy population controls to establish its rare frequency. The impact of mutation on biophysical characteristics of the protein was further studied by mapping it in 3D structure. Furthermore, LC-MS tandem mass spectrophotometry based untargeted metabolomic profiling was done to study the fluctuations in plasma metabolites relevant to disease causative mutations and kidney damage. We identified a very rare homozygous c.631G > A (p.Val211Met) pathogenic mutation in RMND1 gene in the proband, which is inherited in an autosomal recessive fashion. This gene is involved in the mitochondrial translational pathways and contribute in mitochondrial energy metabolism. The p.Val211Met mutation is found to disturb the structural orientation (RMSD is -2.95 Å) and stability (ΔΔG is -0.552 Kcal/mol) of the RMND1 protein. Plasma metabolomics analysis revealed the aberrant accumulation of metabolites connected to lipid and amino acid metabolism pathways. Of these metabolites, pathway networking has discovered ceramide, a metabolite of sphingolipids, which plays a role in different signaling cascades including mitochondrial membrane biosynthesis, is highly elevated in this patient. This study suggests that genetic defects in RMND1 gene alters the mitochondrial energy metabolism leading to the accumulation of ceramide, and subsequently promote dysregulated apoptosis and tissue necrosis in kidneys.

Genetic Mosaicism in Calmodulinopathy.

BACKGROUND: CaM (calmodulin) mutations are associated with congenital arrhythmia susceptibility (calmodulinopathy) and are most often de novo. In this report, we sought to broaden the genotype-phenotype spectrum of calmodulinopathies with 2 novel calmodulin mutations and to investigate mosaicism in 2 affected families. METHODS: CaM mutations were identified in 4 independent cases by DNA sequencing. Biochemical and electrophysiological studies were performed to determine functional consequences of each mutation. RESULTS: Genetic studies identified 2 novel CaM variants (CALM3-E141K in 2 cases; CALM1-E141V) and one previously reported CaM pathogenic variant (CALM3-D130G) among 4 probands with shared clinical features of prolonged QTc interval (range 505-725 ms) and documented ventricular arrhythmia. A fatal outcome occurred for 2 of the cases. The parents of all probands were asymptomatic with normal QTc duration. However, 2 of the families had multiple affected offspring or multiple occurrences of intrauterine fetal demise. The mother from the family with recurrent intrauterine fetal demise exhibited the CALM3-E141K mutant allele in 25% of next-generation sequencing reads indicating somatic mosaicism, whereas CALM3-D130G was present in 6% of captured molecules of the paternal DNA sample, also indicating mosaicism. Two novel mutations (E141K and E141V) impaired Ca2+ binding affinity to the C-domain of CaM. Human-induced pluripotent stem cell-derived cardiomyocytes overexpressing mutant or wild-type CaM showed that both mutants impaired Ca2+-dependent inactivation of L-type Ca2+ channels and prolonged action potential duration. CONCLUSIONS: We report 2 families with somatic mosaicism associated with arrhythmogenic calmodulinopathy, and demonstrate dysregulation of L-type Ca2+ channels by 2 novel CaM mutations affecting the same residue. Parental mosaicism should be suspected in families with unexplained fetal arrhythmia or fetal demise combined with a documented CaM mutation.

Modulation of doxorubicin-induced expression of the multidrug resistance gene in breast cancer cells by diltiazem and protection against cardiotoxicity in experimental animals.

BACKGROUND: Doxorubicin (DOX) is one of the most important anticancer agents used in treating breast cancer. However, chronic cardiotoxicity and multidrug resistance limit the chemotherapeutic use of DOX. METHODS: This study aimed to evaluate the capability of calcium channel blocker diltiazem (DIL) to reverse DOX resistance in breast cancer MCF-7 cells and to confer protection against DOX-induced cardiotoxicity in Wistar rats. For this purpose, we explored the effects of DOX on cell cycle phase distribution and expression of ABCB1, FOXO3a, and p53 genes in the presence and absence of DIL (20 µg/ml) and studied the ability of DIL to prevent DOX-induced cardiotoxicity after a single injection of DOX (15 mg/kg) in male Wister rats. RESULTS: We found that compared with DOX alone treatment, DIL + DOX treatment down regulated the ABCB1 gene expression by > fourfold but up regulated the FOXO3a and p53 genes expression by 1.5 fold. DIL treatment conferred protection against DOX-induced cardiotoxicity, as indicated by a decrease in the levels of the cardiac enzyme creatine kinase MB and malondialdehyde and an increase in the total antioxidant capacity and glutathione peroxidase levels. These biochemical results were further confirmed by the histopathological investigation of cardiac cells, which showed normal cardiac cells with central vesicular nuclei and prevention of DOX-induced disruption of normal cardiac architecture in the DIL to DOX group. CONCLUSIONS: Taken together, our results indicate that DIL treatment can reverse the resistance of breast cancer cells to the therapeutic effects of DOX and can protect against DOX-induced cardiotoxicity in rats.

A Novel Homozygous Frameshift Mutation in CCN6 Causing Progressive Pseudorheumatoid Dysplasia (PPRD) in a Consanguineous Yemeni Family.

Background: Progressive pseudorheumatoid dysplasia (PPRD) inherited in an autosomal recessive fashion, is a disabling disease, characterized by platyspondyly, irregularities of the vertebral bodies, narrowing of the intervertebral discs and intraarticular spaces, widening of the epiphysis-metaphysis, polyarthralgia, multiple joint contractures, and disproportionate short stature. A number of studies have been performed on this deformity in various populations around the globe, including the Arab population. Mutations in CCN6, located on 6q22, are reported to cause this anomaly. Case Presentation: The present study describes the investigation of a consanguineous family of Yemeni origin. Clinical examination of the patient revealed short stature with progressive skeletal abnormalities, stiffness and enlargement of small joints of the hands along with restriction of movements of proximal interphalangeal (PIP) and distal interphalangeal (DIP) joints with weakness and gait disturbance. Sanger sequencing revealed a novel homozygous frameshift deletion mutation (c.746delT; p.Val249Glyfs*10) in CCN6 which may lead to NMD (Nonsense mediated decay). This mutation expands the spectrum of pathogenic variants in CCN6 causing PPRD.

Molecular Screening of VAX1 Gene Polymorphisms Uncovered the Genetic Heterogeneity of Nonsyndromic Orofacial Cleft Among Saudi Arabian Patients.

OBJECTIVE: Nonsyndromic orofacial cleft (NSOFC) including cleft lip with or without cleft palate (CL±P) and cleft palate (CP) are multifactorial developmental disorders with both genetic and environmental etiological factors. In this study we investigated the association between CL±P and CP, and two polymorphisms previously determined using genome-wide association studies, as well as the association between consanguinity and CL±P and CP. METHODS: DNA was extracted from saliva specimens from 171 triads consisting of affected individuals and their parents, as well as 189 control triads (matched for age, gender, and location) that were recruited from 11 referral hospitals in Saudi Arabia. Two polymorphisms, rs4752028 and rs7078160, located in the VAX1 gene were genotyped using real-time polymerase chain reaction. A transmission disequilibrium test was carried out using the Family-Based Association Test and PLINK (genetic tool-set) to measure the parent-of-origin effect. RESULTS: Significant differences were found between affected individuals and the control group. In the case of the rs4752028 risk allele in cleft, the phenotypes were: CL±P (fathers: odds ratio [OR] 2.16 [95% CI 1.38-3.4]; mothers: OR 2.39 [95% CI 1.53-3.71]; and infants: OR 2.77 [95% CI 1.77-4.34]) and CP (fathers: OR 2.24 [95% CI 1.15-4.36] and infants: OR 2.43 [95% CI 1.25-4.7]). For CL±P and the rs7078160 risk allele, the phenotypes were: (fathers: OR 1.7 [95% CI 1.05-2.86]; mothers: OR 2.43 [95% CI 1.49-3.97]; and infants: OR 2.34 [95% CI 1.44-3.81]). In terms of consanguinity, we found significant association between consanguinity and the rs4752028 polymorphism minor allele among CL±P compared with controls (p = 0.001). CONCLUSION: This is the first study to find a relationship between these two loci on 10q25 (rs4752028 and rs7078160) and NSOFC in a population with high levels of consanguinity.

Detection of Secondary Metabolites as Biomarkers for the Early Diagnosis and Prevention of Type 2 Diabetes.

BACKGROUND: Type 2 diabetes, or T2D, is a metabolic disease that results in insulin resistance. In the present study, we hypothesize that metabolomic analysis in blood samples of T2D patients sharing the same ethnic background can recover new metabolic biomarkers and pathways that elucidate early diagnosis and predict the incidence of T2D. METHODS: The study included 34 T2D patients and 33 healthy volunteers recruited between the years 2012 and 2013; the secondary metabolites were extracted from blood samples and analyzed using HPLC. RESULTS: Principal coordinate analysis and hierarchical clustering patterns for the uncharacterized negatively and positively charged metabolites indicated that samples from healthy individuals and T2D patients were largely separated with only a few exceptions. The inspection of the top 10% secondary metabolites indicated an increase in fucose, tryptophan and choline levels in the T2D patients, while there was a reduction in carnitine, homoserine, allothreonine, serine and betaine as compared to healthy individuals. These metabolites participate mainly in three cross-talking pathways, namely "glucagon signaling", "glycine, serine and threonine" and "bile secretion". Reduced level of carnitine in T2D patients is known to participate in the impaired insulin-stimulated glucose utilization, while reduced betaine level in T2D patients is known as a common feature of this metabolic syndrome and can result in the reduced glycine production and the occurrence of insulin resistance. However, reduced levels of serine, homoserine and allothrionine, substrates for glycine production, indicate the depletion of glycine, thus possibly impair insulin sensitivity in T2D patients of the present study. CONCLUSION: We introduce serine, homoserine and allothrionine as new potential biomarkers of T2D.

Autosomal recessive long QT syndrome, type 1 in eight families from Saudi Arabia.

BACKGROUND: One of the most common primary cardiac arrhythmia syndromes is autosomal dominant long QT syndrome, type 1 (LQT1), chiefly caused by mono-allelic mutations in the KCNQ1 gene. Bi-allelic mutations in the KCNQ1 gene are causal to Jervell and Lange-Nielsen syndrome (JLNS), characterized by severe and early-onset arrhythmias with prolonged QTc interval on surface ECG and sensorineural deafness. Occasionally, bi-allelic mutations in KCNQ1 are also found in patients without any deafness, referred to as autosomal recessive long QT syndrome, type 1 (AR LQT1). METHODS: We used Sanger sequencing to detect the pathogenic mutations in KCNQ1 gene in eight families from Saudi Arabia with autosomal recessive LQT1. RESULTS: We have detected pathogenic mutations in all eight families, two of the mutations are founder mutations, which are c.387-5T>A and p.Val172Met/p.Arg293Cys (in cis). QTc and cardiac phenotype was found to be pronounced in all the probands comparable to the cardiac phenotype in JLNS patients. Heterozygous carriers for these mutations did not exhibit any clinical phenotype, but a significant number of them have sinus bradycardia. CONCLUSION: To the best of our knowledge, this is the first description of a large series of patients with familial autosomal recessive LQT, type 1. These mutations could be used for targeted screening in cardiac arrhythmia patients in Saudi Arabia and in people of Arabic ancestry.

Attitude toward Prenatal Testing and Termination of Pregnancy among Health Professionals and Medical Students in Saudi Arabia.

This study was aimed at assessing the attitude of health care professionals in Jeddah city toward prenatal diagnosis (PND) and termination of pregnancy (TOP). A cross-sectional study was conducted, and the participants completed a self-administered questionnaire. Approximately 82% of participants showed a consistent trend of accepting PND when appropriate, and 47.5% of the respondents were in favor of TOP if the fetus had a severe disease. Compared with men (69.3%), a significantly greater number of women (88%) accepted to have PND. The most acceptable prenatal diagnostic tests in the study were invasive techniques as most of the participants thought that noninvasive tests were nonspecific.

Ramadan fasting in Saudi Arabia is associated with altered expression of CLOCK, DUSP and IL-1alpha genes, as well as changes in cardiometabolic risk factors.

BACKGROUND: During the fasting month of Ramadan, practicing Saudis develop severe disturbances in sleeping and feeding patterns. Concomitantly, cortisol circadian rhythm is abolished, diurnal cortisol levels are elevated and circulating levels of several adipokines are altered favouring insulin resistance. AIM: To examine changes in the expression of CLOCK and glucocorticoid-controlled genes, such as DUSP1 and IL-1α in Saudi adults before and during Ramadan, and to investigate possible associations with selected cardiometabolic risk factors. METHODS: Healthy young volunteers (5 females, 18 males; mean age +SEM = 23.2 +1.2 years) were evaluated before Ramadan and two weeks into it. Blood samples were collected at 9 am (±1 hour) and twelve hours later for determination of serum lipid profile, high sensitivity CRP (hsCRP), and adiponectin. The expression of CLOCK, DUSP1 and IL-1α was evaluated in circulating leukocytes. RESULTS: Mean levels of GGT and morning adiponectin decreased, while those of LDL-c/ HDL-c and atherogenic index (AI) increased significantly in Ramadan compared to Shabaan. There was no significant difference between morning and evening adiponectin during Ramadan, while the diurnal rhythm of hsCRP was lost. CLOCK gene expression mean was significantly higher in morning than in evening during Shabaan. Mean morning and evening DUSP1 mRNA levels showed significant increase during Ramadan compared to Shabaan, however, its diurnal rhythm was maintained. Morning IL-1α mRNA expression remained significantly higher than in the evening during Ramadan, but was markedly decreased compared to Shabaan. DISCUSSION: Ramadan fasting in Saudi Arabia is associated with improvements in some cardiometabolic risk factors, such as circulating GGT and hsCRP and leukocyte expression of IL-1α mRNA, suggesting that intermittent fasting might have a beneficial component. These benefits may be offset by the previously reported dysregulation in the circadian rhythm, excess glucocorticoid levels and action, and insulin resistance, explaining increased prevalence of cardiometabolic disorders and type 2 diabetes mellitus.